Certain engineered nanomaterials (ENM) have been shown to cause significant lung pathological changes in animal models raising concern that human health effects will emerge with increasing use and exposure. However, a mechanistic predictive model based on physical and surface properties of ENM has not been established to aid in protecting human health. Other work in our laboratory has shown an association between in vitro NLRP3 inflammasome (NLRP3) activation and in vivo inflammation. In preliminary data, we have shown that there may be a correlation between the ability of nickel associated MWCNT (Ni-MWCNT) to activate NLRP3 in vitro using primary alveolar macrophages (AM) or THP-1 cells with lung inflammation and pathology. Therefore, we propose that in vitro activation of NLRP3 appears to be a reliable predictor of lung inflammation. The basis for the distinction between bioactive and benign ENM is most likely associated with the ability of ENM to be phagocytosed and/or ability to disrupt lysosomes causing cathepsin B release. We hypothesize that the in vivo inflammatory potential of ENM correlates well with lysosomal disrupting activity and NLRP3 activation. Based on preliminary data we propose that the inflammatory potential of MWCNT will be dependent on the amount of associated nickel. Therefore, our aims are: 1. Determine the relationship between NLRP3 inflammasome activity in AM and lung pathology in mice following exposure to defined and well- characterized Ni-MWCNT. 2. Determine the importance of Ni-MWCNT surface properties (amount of Nickel and surface charge) to cause cytotoxicity and degree of activation of the NLRP3 inflammasome using THP-1 cells. Finally, we will establish that internalization of MWCNT-Ni followed by lysosomal membrane disruption and Cathepsin B release is the initiating event in activation of the NLRP3 inflammasome. Thus, information from this study will be important in determining characteristics of safe ENM and establish mechanisms of action and may lead to an in vitro screening platform using THP-1 cells. PUBLIC HEALTH RELEVANCE: These studies will establish the potential of nanomaterials to disrupt lysosomal membranes and activate the NRLP3 inflammasome to cause in vivo inflammation and pathology. Furthermore that the assays performed in the proposal may serve as tools for high throughput screening of ENM. !